Study On The Improvement Of The Production Of Spinosad By Rejuvenation And Coculture

Spinosad,which was the production of aerobic fermentation of Saccharopolyspora spinosa,was a macrolide broad-spectrum insecticide.In this dissertation,the methods of rejuvenation and the coculture were used to influence on the output of spinosad.The dissertation included the study of the effect of rejuvenation methods on the output of spinosad,the screening and the identification of coculture strains and the study of the mechanism of coculture.The purification rejuvenation,the elimination rejuvenation and the larvae rejuvenation of one strain of Saccharopolyspora spinosa 54-ssy whose production was declined were studied.The strain morphology,the output of spinosad and the toxicity showed that the elimination rejuvenation could significantly increase the toxicity and the output of spinosad.Through the resistant rejuvenation of erythromycin,the lethality rate of the strain reached 100%and the yield of spinosad increased by 10 times.Above all,the elimination rejuvenation was the best method of rejuvenation.Some strains were isolated from marine and soil,others were saved in laboratory.All of them were cocultured with Saccharopolyspora spinosa.The results of the output and the toxicity of spinosad showed that strain SY could improve the output of spinosad.The evidence of the strain morphology,16S rDNA,Biolog and G+C mol%proved that strain SY was Rhodococcus.The strain SY was named Rhodococcus sp.SY.The optimization of the fermentation medium and conditions was carried out.The most suitable medium for coculture consisted of glucose 6.45%,maltose 1.0%,cottonseed flour 2.5%,peptone 1.2%,yest extract 1.2%.An inoculation of 1%Rhodococcus at the 108 h and an 72 h coculture of the Saccharopot yspora spinosa fermentation leaded to the highest output of spinosad.The theories of Horizontal Gene Transfer,complementary metabolism and inducible expression were used to study the mechanism of coculture.The results suggested that the mycolic acid present in the outer layer of cell is important for inducing spinosad production,and the living cell play a critical role in the inducing activity.